1. Field of the Invention
The present invention relates to a culture method, and more particularly, to a method for decreasing feed conversion rate.
2. Description of the Related Art
With the rapid increase in world population and decrease in arable land area, the available feed resources get increasingly short, and improvement of output per unit of area and increase of additional nutritional value are tendencies in feed production in the world.
As for the fish industry, because the fish harvest obtained from fishing from the sea is almost saturated, and the demand for fish products on the animal protein market daily increases, it is necessary to supplement the shortage in product supply from fishing industry with aquaculture.
Epinephelus spp. is a kind of warm-water fish belonging to Epinephelus, Epinephelinae, Serranida, Perciforms, Osteichthyes, is widely distributed in tropical and subtropical waters around the world, and is generally acknowledged as the most important economic fish species in the Asia and Pacific regions by the culture industry. In Taiwan, the fish culturinges are mainly Epinephelus malabaricus, Epinephelus coioides, Epinephelus lanceolatus and Epinephelus fario. Generally, the Epinephelus spp. must be cultured for at least 8 to 12 months before marketing; however, the final yield is not high. In addition to earlier mortality of young fish due to viral and bacterial infection, for example, weather factors, water pollution problems, selection of feed organisms and management during culturing all will increase risk to the practitioner. Therefore, how to improve the development rate of fish culturinges, shorten the culture time before marketing, and decrease the feed conversion rate (weight of feed fed/gain of body weight of fish body) and the potential risks during culturing has become a research direction worthy of hard work.
Secreted Protein Acidic and Rich in Cysteine (SPARC), also referred to as osteonectin or BM40 with a protein size of about 35-45 kDa and mainly distributed in the extracellular matrix (ECM), is a matricelluar protein, and is also a multi-functional glycoprotein having a property of binding to the ECM or a cell.
Although the SPARC is mainly distributed in the ECM; however, the function of the SPARC is different from a common structural protein in the matrix which is responsible for the structure constitution and supporting of a cell; on the contrary, the SPARC specially servers as a bridge connecting the cell and the ECM, and is a regulatory protein capable of regulating the production, storage and accumulation of several types of cellular matrix protein (Bradshaw and Sage, 2001, J Clin Invest 107; 1049-1054; Lane and Sage, 1994, FASEB J 8; 163-173). However, the SPARC is mainly expressed during embryo development, has great influence on cell differentiation, calcification and generation of tissue, bone development, morphology, and organ development. The expression level of this protein in an adult organism trends to be lowered, because obvious expression occurs mainly in a repair and reorganization process of injured skin or tissue (Lane and Sage, 1994).
The SPARC can be mainly divided into three functional domains (Hohenester et al., 1997, EMBO J 16; 3778-3786): 1. acidic domain at a N terminal of the protein, which is capable of bonding with 5-8 calcium ions with weak affinity and has functions of inhibiting cell extension and regulating production of ECM; 2. Follistatin like domain, which includes multiple Cysteines, has function similar to follistatin, and is capable of inhibiting cell proliferation, and further includes another specific sequence, that is, a copper ion binding sequence (K) GHK (Iruela-Arispe et al., 1995, Mol Biol Cell 6; 327-343), and has vascular proliferation promoting function; and 3. extracellular calcium ion binding domain (EC domain), which is a C-terminal of SPARC, has two EF-hand motifs, is capable of binding with calcium ions with high affinity, and is capable of binding with a cell or some collagens, and further has effect of inhibiting cell proliferation and extension (Maurer et al., 1995, J Mol Biol 253; 347-357).
Currently, some results are got through researches and discussions in biological functions of SPARC, and several properties are found through observation of in vitro endothelial cell culture of mouse: (I) expression of SPARC can inhibit the extension of cell morphology to make a cell nearly round (Murphy-Ullrich et al., 1991, J Cell Biol 115; 1127-1136); regulating the composition of ECM and expression of some ECM proteins, to adjust the binding of a cell with an extracellular matrix to change the cell morphology and migration ability (Hasselaar et al., 1991, J Biol Chem 266; 13178-13184; and Tremble et al., 1993, J Cell Biol 121; 1433-1444); (II) inhibiting the progression of cell cycle, to arrest the cycle in mid-G1 phase, and thus having cell growth inhibiting function (Yan and Sage, 1999, J Histochem Cytochem 47; 1495-1506); (III) it was found in researches of vascular cell culture that endothelial cells have a vascular proliferation promoting property (Funk and Sage, 1991, Proc. Natl. Acad. Sci. USA 88; 2648-2652). In vivo, it is found that mouse with SPARC gene knocked down is afflicted with cataract no long after being born, and thus it is postulated that SPARC is an essential protein for development of eye lens (Yan and Sage, 1999). It is also found through researches that in mice with sparc gene knocked down, abnormal changes in skin structure, thickened fatty layer, increases in volume and number of lipocyte, and rise of concentration of leptin secreted by lipocytes in the blood are observed, and thus it is postulated that the SPARC protein has correlation to in vivo regulation of the changes of the adipose amount in the body (Bradshaw et al., 2003, Proc. Natl. Acad. Sci. USA 100; 6045-6050).
Myostatin, also referred to as growth and differentiation factor-8 (GDF-8), is a member in transforming growth factor TGF-β family. Myostatin may negatively regulate the growth and differentiation of muscle. It can be found through researches in bovine and human bodies that when the function of myostatin is lost, because the negative regulation of the growth and differentiation of muscle cannot effect, a phenotype of muscle increasing occurs (McPherron and Lee, 1997, Proc. Natl. Acad. Sci. USA 94, 12457-12461; and Schuelke et al, 2004, The New England Journal of Medicine 350, 2682-2688). In the researches with mammal, myostatin in mice is knocked down (McPherron and Lee, Proc. Natl. Acad. Sci. USA 94; 12457-12461), high-level expression of myostatin inhibiting protein (for example, follistatin) in the muscle of rice or disabling the function the receptor protein ActRIIB essential for signal transmission (Lee and McPherron, 2001, Proc. Natl. Acad. Sci. USA 98; 9306-9311) will cause the occurrence of muscle increasing. In the researches with bony fishes, antisense morpholino knock-down technology is used to inhibit mRNA of myostatin-1 in the embryo of zebra fish, so as to accelerate the embryo growth (Amali et al, 2004, Developmental Dynamics 229; 847-856); or RNAi technology is used to silence myostatin-1 in zebra fish, and thus features of giant zebra fish occur (Acosta et al, 2005, Journal of Biotechnology 119; 324-331). The muscle cells are further researched with tissue slice, and it is found that hypertrophy of the volume and hyperplasia of the number of the muscle cells may be causes causing muscling.
On the other hand, it is pointed out in in-vitro cell line researches that, if myostatin is expressed in C2C12 myoblast line at high level, the proliferation of myoblast can be inhibited, to arrest the cell in G1 to S phases in the cell cycle (Thomas et al, 2000, The Journal of Biological Chemistry 275; 40235-40243). On the other hand, if myostatin is expressed in muscle astrocyte at high level, it is found that myostatin may inhibit the activation and self-renewal of muscle astrocyte (McCroskery, 2003, The Journal of Cell Biology 162; 1135-1147), which also proves that myostatin does play a role of negative regulation in the development of muscle. In addition to the regulation in muscle cells, it is pointed out in other literatures that myostatin has the function of inhibiting adipogenesis (Rebbapragada et al, 2003, Molecular and Cellular Biology 23; 7230-7242), and it is also pointed out in researches that myostatin has the function of inhibiting apoptosis of myoblasts (Rios et al, 2001, Biochemical and Biophysical Research Communications 280; 561-566).
However, in researches of myostatin and SPARC, there is no reports about the feed conversion rate (weight of feed fed/gain of body weight), which is also critical for cost control in culture industry, in addition to muscle gain. Therefore, a method for decreasing the feed conversion rate is still needed to be developed in the industry, so as to improve the development rate of culturing.